The field of self-assembling peptides (SAP) is an emerging technology area whose rapid growth is fueled by the potential for applications in tissue engineering, biotechnology, nano- and regenerative medicine, nanoengineering, synthetic biology, and optics and electronics. The peptides self assemble to create nanostructures that can promote biological activity (such as nerve regeneration, bone growth, and angiogenesis); enhance active delivery or release; modify liquid rheological properties; and coat or modify surfaces. Additionally, they can create even larger micro- and macrostructures such as membranes, fibrils, fibers and potentially even fabrics.
Molecular self-assembly may be defined as the spontaneous and reversible organization of molecular units into ordered structures, driven by non-covalent interactions (Whitesides et al.; Science 1991, 254, 1312-1319). The spontaneous nature of self-assembly processes requires a lowering of the free energy of the system and therefore the formation of the nanostructure from monomer units is commonly associated with a critical concentration of monomer units for self-assembly (Aggeli et al.; PNAS 2001, 98, 11857-11862). Prior to the critical concentration, the chemical potential of the monomer species rises rapidly as monomer is added to the system until the critical concentration where the nanostructure starts to form. Above the critical concentration all further monomer added to the system will add to and enlarge the nanostructures with the chemical potential of the monomer species effectively fixed at a set value.
There are multiple secondary structures that are known to form, including extended beta strands (cross-beta structure) that form tapes, tubes and fibers driven by hydrogen bonding parallel to the tape axis, and alpha-helices which form triple-helix collagen-like structures and coiled-coil bundles through side-chain interactions perpendicular to the fiber axis. Of these beta-sheet self-assembly is acknowledged to be the simplest form, where self-assembly is typically driven by a combination of hydrogen bonds, complimentary charge, aromatic and Van der Waals interactions between the monomer peptide residues. Each residue can be estimated to contribute approximately 2-3 kT of free energy to the system, where 1 kT is equal to the amount of thermal energy that is present in a chemical system at room temperature and pressure. Simplistically, therefore any structure which has more than 1 kT in free energy is likely to form thermodynamically-stable structures and the greater this value the more stable and long-lived these structures will be in the chemical system. As a result short peptides are observed to form very large stable self-assembled structures in solution, at interfaces and on surfaces. Remarkably, even dipeptides containing only two amino acids have been shown to self-assemble into beta-sheet peptide tapes that are hundreds of microns in length (Reches and Gazit; Curr. Nanoscience 2006, 2, 105-111). More generally, self assembling oligopeptide structures fall into two major classifications: those consisting of sequences of hydrophobic and hydrophilic amino acids which result in amphiphilic properties (an amphiphilic peptide), and those where the amphiphilic properties are the result of modification of the peptide with a hydrophobic group (a peptide amphiphile).
The work of Aggeli, Zhang and Stupp among others has allowed a rational set of design criteria to be defined for self-assembling beta-sheet tape forming peptides and peptide amphiphiles, which allows the skilled practitioner to design and create new self-assembling peptide sequences with reasonable confidence. Based on these seminal studies, it can be seen that in general, increasing the number of residues will decrease the critical concentration as self-assembly becomes more favorable through the increased number of complimentary interactions between the monomer peptides. Solvent conditions including pH, salt concentration, dielectric and temperature have been shown to have significant influence upon the self-assembly behavior dependent on the primary sequence. For oligopeptides containing several charged residues, where there is significant coulombic repulsion between the monomeric peptides, raising the salt concentration will more efficiently screen and neutralize like-charges between monomers and lower the critical aggregation concentration. This is observed with the behavior of the RADA peptides where contact of monomeric solution with higher salt concentration causes instant gelation (Ellis-Behnke et. al.; Nanomedicine: Nanotechnology, Biology and Medicine 2006, 2 (4), 207-215). Similarly, pH has significant influence on the self-assembly of oligopeptides containing charged residues, where in general for self-assembly of anionic peptides when the pH is lowered below the pK of the anionic residues, the net charge on the monomeric peptides will be neutralized and the critical aggregation concentration will be lowered. Similarly for self-assembly of cationic peptides when the pH is raised above the pK value of the cationic residues, the net charge is neutralized and the critical aggregation concentration will be lowered (Aggeli et al.; Angew. Chem. Int. Ed. 2003, 42, 5603-5606). Commonly most self-assembling peptides would contain a mixture of anionic and cationic charged residues in which case the specific response to pH and salt concentration will be sequence dependent. Additionally it should be recognized that this responsive behavior can be engineered by designing specific sequences that spontaneously self-assemble in the presence of specific environmental conditions such as an increase in salt concentration due to contact with body fluids or sweat insults or change in pH due to contact with keratinous substrates such as skin or hair.
Another significant class of self assembling oligopeptides is the group of peptide amphiphiles, wherein the peptide has been functionalized with an alkyl group (C12-C22). (Hamley; Soft Matter, 2011, 7, 4122-4138) Examples of this class of materials have been described to self-assemble in aqueous media, with or without the presence of polymeric compounds, to give nanofibers that can further aggregate to form strong and flexible macrofibers that can be multiple centimeters in length (Capito, et. al.; Science 2008, 319, 1812-1816). These nanofibers can be induced by changing (1) the salt concentration, (2) the pH, or (3) the moisture content of the system. They can be used for various medical applications such as drug delivery, medical diagnostic components, tissue, cartilage, enamel or nerve regeneration, cell growth promotion, and wound healing. Temperature effects have also been extensively explored within the literature and recently were shown by Stupp and coworkers to be of significant importance for affecting subtle rearrangement of self-assembled peptide structures producing significant changes to the rheological behavior and stability of the peptide structures (Zhang et al., Nature Materials 2010, 9, 594-601). A heat pre-treatment of the peptide amphiphile solution was used followed by equilibration to lower temperature. This promoted a reorganisation of the self-assembled structure and allowed for formation of aligned nanofiber bundles, creating macrofibers on the order of centimeters long, which were stabilized by exposure to calcium salt solutions.
Similarly studies on collagen mimics and coiled coil peptide sequences have shown that environmental conditions can be used to control the self-assembly process enabling a large range of applications where responsive behavior is required or desired (Fletcher et al., Soft Matter 2011, 7, 10210; O'Leary et al.; Nature Chemistry 2011, 3, 821-828).
These examples demonstrate how self-assembly behavior can be engineered for specific applications based upon rational design of the primary sequence which in turn determines the macroscopic physical properties of the self-assembled materials. However predicting responsive behavior or the exact critical aggregation concentration for a given oligopeptide cannot be done purely from a knowledge of the primary sequence. Additionally, the peptide will behave differently as solution conditions of pH, ionic strength, temperature, and co-ingredients vary, which will affect the concentration of oligopeptide required to form self-assembled nanofibers in different compositions to achieve the desired behaviour under different application conditions. It can be said that in general: increasing the number of residues will decrease the critical concentration and self-assembly will become more favorable; raising the salt concentration will more efficiently screen like-charges between monomers and lower the critical concentration; and decreasing the temperature will have a similar effect. Thus for purely thickening a composition, a ten residue peptide would typically be present at a lower concentration than a three residue peptide as the ten residue peptide is likely to be more efficient at increasing the viscosity.
As another example Zhang and Ellis-Behnke have shown that application of RADA16-I (SEQ ID NO: 1) to open wounds can instantly stop bleeding (Haemostasis) through salt-induced peptide self-assembly to form fibrous hydrogel transparent membranes that encapsulate and prevent further blood loss from the wound (Ellis-Behnke et. al. Nanomedicine: Nanotechnology, Biology and Medicine 2006, 2 (4), 207-215). These membranes can be envisioned to provide a variety of other benefits in personal care compositions such as skin protection, active delivery, wrinkle reduction, and wetness protection. Skin actives require penetration into the skin in order to function. The thermodynamic favorability of penetration is critical. For that reason, the concentration of the material in the composition must be near the saturation point for optimum penetration. The most significant cost in producing a composition comprising a skin active is the cost of producing or obtaining the skin active itself. Thus, there is a need in the arts of cosmetic and pharmaceutical compositions applied to the skin to efficiently deliver a skin active to, into, and/or through the skin, which could be enabled by incorporation of self assembling peptides.
Notwithstanding these developments, a need continues to exist to develop compositions, and methods of using those compositions, that provide improved or new benefits via personal care compositions comprising self assembled peptides, for example to improve the delivery of skin or hair benefit agents to the skin or hair, to provide hair and skin surface modification, or to create elongated hair fibers.